Total Protein Extraction Kit (Strong), IC-7901

Product Introduction

This product is a ready-to-use kit for the efficient extraction of total protein from cultured cells and tissues using a strong denaturing lysis buffer system. Complete and efficient protein extraction is a critical prerequisite for accurate and reproducible downstream protein analysis, particularly for proteins that are poorly soluble, membrane-associated, nuclear, or present in protein complexes. Traditional mild lysis buffers may incompletely solubilize these protein populations, leading to selective protein loss and biased representation in subsequent analyses. This kit employs a strong denaturing buffer formulation containing high concentrations of chaotropic agents and detergents that effectively disrupt all cellular compartments, solubilize membrane proteins, dissociate protein complexes, and denature proteases, ensuring comprehensive extraction of the total cellular proteome. The extracted proteins are compatible with SDS-PAGE, Western blotting, and protein quantification assays following appropriate dilution or buffer exchange. This kit is particularly suitable for applications requiring maximal protein recovery, including total proteome analysis, extraction from difficult-to-lyse tissues, and studies of membrane proteins, nuclear proteins, and insoluble protein aggregates.

Product Features

1. Strong Denaturing Lysis Buffer: High-concentration chaotropic agents and detergents ensure complete solubilization of membrane proteins, nuclear proteins, and protein complexes.

2. Comprehensive Proteome Extraction: Recovers total cellular protein including difficult-to-extract proteins that may be underrepresented in mild lysis buffer extracts.

3. Effective Protease Inhibition: Denaturing conditions inactivate endogenous proteases, preserving protein integrity during extraction.

4. Versatile Sample Compatibility: Suitable for cultured cells (suspension and adherent), fresh tissues, and frozen tissues.

5. Ready-to-Use Kit Format: Includes all necessary extraction buffer and protease inhibitor cocktail for a complete workflow.

Specifications

Size: 100 extractions

Sample Types: Cultured cells (suspension and adherent), fresh and frozen tissues

Extraction Type: Total protein (denaturing)

Lysis Buffer: Strong denaturing formulation with chaotropic agents and detergents

Downstream Applications: SDS-PAGE, Western blotting, protein quantification

Storage and Stability

Storage Conditions: Store kit components at 4¡ãC. Protease inhibitor cocktail should be stored at -20¡ãC. Protect from light.

Shelf Life: The product is stable for 12 months from the date of manufacture when stored as directed.

Protocol (For Reference Only)

Important: Add protease inhibitors freshly to the extraction buffer immediately before use. For tissues, pre-chill all equipment and work quickly to minimize proteolysis. The strong lysis buffer denatures proteins; do not use for applications requiring native protein conformation.

1. Buffer Preparation: Immediately before use, add the provided protease inhibitor cocktail to the required volume of Total Protein Extraction Buffer at the recommended ratio. Mix thoroughly and keep on ice.

2. Cell Sample Preparation: For adherent cells, wash cells twice with ice-cold PBS, aspirate all PBS, and add an appropriate volume of ice-cold Total Protein Extraction Buffer directly to the culture vessel (typically 100-200 µL per well of a 6-well plate or 500-1000 µL per T75 flask). Scrape cells thoroughly, transfer the lysate to a microcentrifuge tube, and incubate on ice for 15-30 minutes with intermittent vortexing. For suspension cells, wash twice with ice-cold PBS, pellet cells by centrifugation, add extraction buffer to the pellet, and process as above.

3. Tissue Sample Preparation: For fresh or frozen tissues, mince 20-50 mg of tissue into small pieces and place in a homogenizer. Add 500-1000 µL of ice-cold Total Protein Extraction Buffer and homogenize thoroughly on ice until a uniform suspension is obtained. Transfer the homogenate to a microcentrifuge tube and incubate on ice for 15-30 minutes with intermittent vortexing.

4. Clarification: Centrifuge the lysate at 12,000-14,000 x g for 10-15 minutes at 4¡ãC to pellet insoluble debris. Carefully transfer the supernatant (total protein extract) to a clean pre-chilled tube without disturbing the pellet.

5. Quantification and Storage: Determine protein concentration by BCA or Bradford assay. Due to the high concentration of chaotropic agents and detergents, dilution of the extract or use of a detergent-compatible protein assay is recommended. Use the extract immediately or store at -20¡ãC or -80¡ãC for long-term storage.

Precautions

1. The strong denaturing lysis buffer irreversibly denatures proteins and is not suitable for applications requiring native protein conformation, such as immunoprecipitation or enzyme activity assays.

2. Add protease inhibitors freshly to the extraction buffer immediately before use; do not use buffer pre-mixed with inhibitors after prolonged storage.

3. For tissue samples, ensure thorough homogenization; incomplete homogenization reduces protein yield and represents a major source of variability.

4. Due to the high detergent concentration, use a detergent-compatible protein assay or dilute samples appropriately before protein quantification.

5. For research use only. Not for use in diagnostic or therapeutic procedures.

FAQ (Simplified)

Q1: How does this kit differ from mild lysis buffer kits?

A1: This kit uses a strong denaturing buffer containing high concentrations of chaotropic agents and detergents that completely solubilize all cellular compartments, including membranes, nuclei, and protein complexes. This ensures extraction of difficult-to-solubilize proteins that may be lost with mild lysis buffers, providing a more comprehensive representation of the total proteome.

Q2: Can extracted proteins be used for immunoprecipitation?

A2: No. The strong denaturing conditions irreversibly denature proteins and disrupt antibody-antigen interactions. For immunoprecipitation and other applications requiring native protein conformation, use a mild, non-denaturing lysis buffer kit.

Q3: What protein quantification assay should I use?

A3: Due to high concentrations of detergents and chaotropic agents, a detergent-compatible protein assay (e.g., BCA assay with detergent-compatible modifications, or Bradford assay with appropriate buffer controls) is recommended. Dilute samples in water or assay buffer before quantification if the undiluted extract interferes with the assay.

Q4: Is this kit suitable for phosphorylated protein analysis?

A4: While the denaturing conditions inactivate proteases, the extraction buffer does not contain phosphatase inhibitors by default. For phosphorylated protein analysis, supplement the extraction buffer with a phosphatase inhibitor cocktail in addition to the provided protease inhibitors.

Disclaimer

1. For Research Use Only. Not for use in diagnostic or therapeutic procedures.

2. Due to the variable nature of biological research, optimization of extraction conditions is recommended for specific sample types.

3. This warranty is limited to the replacement of the product. The manufacturer assumes no liability for incidental or consequential damages, including loss of samples or data.

4. Wear appropriate protective clothing and gloves when handling this product.

Ordering Information

Catalog Number: IC-7901

Product Name: Total Protein Extraction Kit (Strong)

Size: 100 extractions

Price: CNY ¥880.00 / USD $88.00 / EUR €105.60 / JPY ¥15840.00

                                                                                                                                                                                                                    2023 Version